CD95/Fas/Apo-1 is a cell surface receptor capable of inducing apoptotic death of human cells. Similar to the physiologic ligand of this receptor, CD95L, agonistic anti-CD95 antibodies may induce apoptosis if binding to CD95 occurs in a multimeric format, e.g., as pentameric IgM or self-aggregating IgG3. Alternatively, anti-CD95 antibodies may be cross-linked by Fc receptors on neighbouring cells or by secondary antibodies to achieve agonistic activity.
Because many tumor cells express CD95, the use of agonistic anti-CD95 antibodies for tumor therapy has been vigorously pursued after initial characterization of prototypic CD95 antibodies. However, it soon became obvious that, at least in its most simple form of applying agonistic anti-CD95 antibodies or recombinant CD95L to patients, this approach fails because many normal cell types express functional CD95 and may be killed by agonistic antibodies.
CD20 is a marker of B-cells involved in many lymphoma and autoimmune diseases, e.g. multiple sclerosis (MS), rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE).
Antibodies directed against the B-cell associated CD20 surface antigen can target normal as well as malignant B cells. They are successfully used for the treatment of B-cell derived leukaemia and lymphoma and antibody mediated autoimmune disease, respectively. Rituximab (trade names Rituxan and MabThera) is a chimeric monoclonal antibody against the protein CD20. Rituximab destroys B cells, and is therefore used to treat diseases which are characterized by excessive numbers of B cells, overactive B cells, or dysfunctional B cells. This includes many lymphomas, leukaemias, transplant rejection, and some autoimmune disorders.
However, rituximab kills CD20-positive cells non-specifically, and was shown to be clinically effective in MS but is compromised by side effects (e.g. Progressive Multifocal Leukoencephalopathy, PML).
It was previously shown that bispecific F(ab′)2 fragments (bs-F(ab′)2) with specificity for CD95 and different target antigens on lymphoma cells, such as CD20 and CD40, induce the apoptosis of cells positive for CD95 and the respective target antigen. Lymphoma cells expressing CD95 but no target antigen were not killed (Jung et al. Cancer Research 61, 1846-1848 (2001)).
Herrmann et al. (Cancer Research 68 (4): 1221-7 (2008) assessed the influence of the antibody format and nature of the target antigen on selective CD95 mediated apoptosis in tumor cells.
US2003/0232049A1 describes a multispecific reagent for selectively stimulating cell surface receptors. Bi-specific antibodies consisting of antigen-binding antibody fragments with a first binding site for a cell surface receptor, such as a death receptor, e.g. CD95, and a second binding site for a target antigen of the same cell, such as CD20 or CD40, are described to kill cancer cells.